Replacing bacterial symbionts with a fungal symbiont… unusual changes in aphids!

Current models of bacterial genome evolution suggest that in small populations a burst of transposable element activity may lead to inactivation of non-essential genes and large deletions, followed by erosion of the pseudogenes resulting in genome reduction. Due to this process, the smallest sequenced cellular genomes are all obligate intracellular symbionts of insects. Interestingly, it seems that the presence of symbionts in small populations within each host reduces the efficacy of the purifying selection increasing the genetic drift that increase the fixation of deleterious mutations.

The evolutionary scenarios seem to be quite different for eukaryotic symbionts since they have different evolutionary patterns of their genomes and it has been reported the gain of mobile genetic elements and intronic sequences resulting in larger genomes.

A recent paper by Kevin J. Vogel and Nancy A. Moran compared the evolution of symbiont analyzing a monophyletic group of aphids within the subfamily Cerataphidinae that have lost the bacterial symbiont common to all other Aphididae (Buchnera aphidicola), which have been replaced by a eukaryotic one, the Yeast-Like Symbiont (YLS). In particular, Vogel and Moran used this system as a model to test the hypothesis that chronically high levels of genetic drift will result in an increase in size of a eukaryotic symbiont genome.

Sequencing of the Yeast-Like Symbiont genome revealed, unlike the obligate bacterial symbionts that lost several genes for DNA repair and recombination, the YLS appears to have fully functional recombinational machinery, including the full suite of genes necessary for meiotic division. Furthermore, the YLS’s genome reveals a diverse suite of metabolic abilities unlike the streamlined metabolism of the obligate bacterial symbionts of insects, though it has lost many genes found in related fungi. At this regards, like Buchnera, the YLS encodes the full biosynthesis pathways for essential amino acids, though it can also produce the non-essential amino acids, which Buchnera mostly receives from the host. The ability of the YLS to produce essential amino acids supports the hypothesis that the YLS has replaced Buchnera’s functional role in these aphids.

The genome of the YLS resulted enriched in introns and presented an elevated rates of amino acid substitution but no burst in mobile DNAs has been observed so that the YLS genome do not support the hypothesis of rampant genome expansion observed in fungi such as Tuber melanosporum. The high gene density and small intergenic spacers suggest that YLS may reside in a range of population size and genome size that allow for expansion of introns but limit the rampant proliferation of mobile genetic elements.

As a whole, the patterns observed in the YLS genome suggest that its symbiotic lifestyle is permissive to intron proliferation and accelerated sequence evolution, though other factors appear to limit its overall genome expansion. This result could suggest the intron and mobile DNA gains occur in different times, but the Buchnera-aphid symbiosis is much older, and is thought to originate at least 200 mya so that the relatively young association of YLS with hosts may not have permitted sufficient time to allow for genome expansion.

Vogel KJ, & Moran NA (2013). Functional and evolutionary analysis of the genome of an obligate fungal symbiont. Genome biology and evolution PMID: 23563967

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Make your choice.. of plants!

Aphids harbour several obligate and facultative bacterial symbionts that have important effects on their life. Several surveys of secondary symbionts clearly show that particular species are strongly associated with aphids feeding on certain food plants. For instance, most pea aphid clones feeding on clover Trifolium sp. harbour Regiella insecticola, while those feeding on Medicago usually have Hamiltonella defensa.

How can we explain such a difference? The most intriguing hypothesis is that these patterns reflect a role of these symbionts in the host plant use. However, they may also be present in view of factors correlated with host plant use or simple historical contingency. So the question is: Can symbiont drive the choice of the plant by aphids or they simply change in view of the plants where aphids live?

Several studies tried to distinguish between these explanations furnishing controversial scenarios. Tsuchida et al. (2004) removed R. insecticola from a clover-associated pea aphid clone using antibiotics and found that performance on Trifolium, but not Vicia, was negatively affected. In the same year, Leonardo repeated the same experimental plan but without finding any fitness effects of removing R. insecticola from two clones of aphid specialized on Trifolium. With a different approach, based on the artificial introduction of R. insecticola into five symbiont-free clones not previously associated with clover, no effect on performance of aphids on Trifolium have been observed by Ferrari et al. (2007). These results, as a whole, suggested that symbionts may be involved in the plant choice but not alone. Probably, interactions between aphids and plants involve the genotype of either the host or symbiont and both can influence host plant use.

Ferrari and Godfray here reported a further set of experiments where they evaluated the fitness consequences of introducing different strains of the symbiont Hamiltonella defensa into three aphid clones (that naturally lack symbionts) collected on Lathyrus pratensis and of removing symbionts from 20 natural aphid–bacterial associations. Ferrari and Godfray reported that: “Infection decreased fitness on Lathyrus but not on Vicia faba, a plant on which most pea aphids readily feed. This may explain the unusually low prevalence of symbionts in aphids collected on Lathyrus. There was no effect of presence of symbiont on performance of the aphids on the host plants of the clones from which the H. defensa strains were isolated. Removing the symbiont from natural aphid–bacterial associations led to an average approximate 20 per cent reduction in fecundity, both on the natural host plant and on V. faba, suggesting general rather than plant-species-specific effects of the symbiont. Throughout, we find significant genetic variation among aphid clones”.

Can you have now a better scenario? As a whole, the results provide no evidence that secondary symbionts have a major direct role in facilitating aphid utilization of particular host plant species, but only the aphid genome seem to have a pivotal role in the plant choice. At present we have a reply, but further experiments on different aphid species are welcome!

References

McLean, A., van Asch, M., Ferrari, J., & Godfray, H. (2010). Effects of bacterial secondary symbionts on host plant use in pea aphids Proceedings of the Royal Society B: Biological Sciences, 278 (1706), 760-766 DOI: 10.1098/rspb.2010.1654
Tsuchida, T. (2004). Host Plant Specialization Governed by Facultative Symbiont Science, 303 (5666), 1989-1989 DOI: 10.1126/science.1094611
Leonardo, T. (2004). Removal of a specialization-associated symbiont does not affect aphid fitness Ecology Letters, 7 (6), 461-468 DOI: 10.1111/j.1461-0248.2004.00602.x
Ferrari, J., Scarborough, C., & Godfray, H. (2007). Genetic variation in the effect of a facultative symbiont on host-plant use by pea aphids Oecologia, 153 (2), 323-329 DOI: 10.1007/s00442-007-0730-2

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Insects? Not just one genome-one organism

In the last day I read with great interest the intriguing review entitled “A symbiotic view of life: we have never been individuals” written by  Scott F. Gilbert, Jan Sapp and Alfred I. Tauber and published in The Quarterly Review of Biology.

Due to their parthenogenetic reproduction aphids are generally considered a sort of clone so that each individual is identical to the others in the population. According to this suggestion, several Authors refereed to aphids as a single genome species. Actually, as well stated by Gilbert et al, the one-genome/one-organism doctrine of classical genetics has been eclipsed by recent studies on symbiosis. In particular, aphid microbial symbionts form a second type of genome and genetic inheritance (Moran 2007; Gilbert 2011). As frequently suggested, insects may acquire their symbionts vertically though the maternal germline as well as horizontally from the environment (such as during feeding). In particular, in aphids, symbiotic bacteria provide selectable allelic variation (thermotolerance, color, parasitoid resistance) that enables some hosts to persist better under different environmental conditions (Dunbar et al. 2007; Tsuchida et al. 2010).

A well-studied example is the pea aphid, Acyrthosiphon pisum since variants of its symbiont Buchnera provide the aphid with thermotolerance, even if at the expense of fecundity at normal temperatures; Dunbar et al. 2007). The second bacterial symbiont Rickettsiella is responsible for color change, turning genetically red aphids into green through the synthesis of quinones (Tsuchida et al. 2010). Furthermore, variants of Hamiltonella symbionts provide immunity against parasitoid wasp infection (Oliver et al. 2009). Interestingly, in the last case, the protective role of Hamiltonella is due to the incorporation of a specific lysogenic bacteriophage within the bacterial genome. The aphids are therefore infected by Hamiltonella that must be infected by phage APSE-3. As Oliver et al. (2009) wrote, “In our system, the evolutionary interests of phages, bacterial symbionts, and aphids are all aligned against the parasitoid wasp that threatens them all. The phage is implicated in conferring protection to the aphid and thus contributes to the spread and maintenance of H. defensa in natural A. pisum populations” .

However, symbioses are frequently not for free for the hosts and even if aphids have some advantages due to symbionts in the presence of parasitoids having their beneficial protection, in the absence of parasitoid wasps aphids carrying the bacteria with lysogenic phage are not as fecund as those lacking them. Similarly, a trade-off occurs in aphids that carry the thermotolerant genetic variants of Buchnera, meaning that more heat-resistant aphids have less fecundity at milder temperatures than their sisters whose bacteria lack the functional allele for the heat-shock protein.

As Gilber et al reminded at the ned of their review, not all scientists involved in evolution agree about the important role of symbiosis so that, for instance, in the 2009 “Homage to Darwinism” debate held at Oxford University, Richard Dawkins questioned the bringing of symbiosis into evolutionary theory: “Take the standard story for ordinary animals, [where] you’ve got a distribution of animals [and] you’ve got a promontory, or  an  island or something and so you end up with two [geographical] distributions. And then on either side you get different selection pressures, and so one [group] starts to evolve this way, and [the other] one starts to evolve that way, and what’s wrong with that? It’s highly plausible, it’s economical, it’s parsimonious. Why on Earth would you want to drag in symbiogenesis when it’s so unparsimonious and uneconomical?”. As Lynn Margulis replied at that time…  simply because symbiosis exists and it is common in living organisms.

 References

• Dunbar HE, Wilson AC, Ferguson NR, & Moran NA (2007). Aphid thermal tolerance is governed by a point mutation in bacterial symbionts. PLoS biology, 5 (5) PMID: 17425405

• Gilbert S. F. 2011. Symbionts as genetic sources of hereditable variation. pp. 283–293. In Transformations of Lamarckism: from sbtle fluids to molecular biology, edited by S. B. Gissis and E. Jablonka. Cambridge (Massachusetts): MIT Press.

• Moran NA (2007). Symbiosis as an adaptive process and source of phenotypic complexity. Proceedings of the National Academy of Sciences of the United States of America, 104 Suppl 1, 8627-33 PMID: 17494762

• Oliver, K., Degnan, P., Hunter, M., & Moran, N. (2009). Bacteriophages Encode Factors Required for Protection in a Symbiotic Mutualism Science, 325 (5943), 992-994 DOI: 10.1126/science.1174463

• Tsuchida T., Koga R., Horikawa M., Tsunoda T., Maoka T., Matsumoto S., Simon J.-C., Fukatsu T. 2010. Symbiotic bacterium modifies aphid body color. Science 330:1102–1104.

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Aphids and their ecological immunity

Bacteria commonly interact with aphids in intimate symbioses, where symbionts increase host fitness (for a review see Russell and Moran, 2006). Interestingly, several evidences suggested that symbiotic bacteria present in the insect gut are involved not only in the degradation of specific substances in the food (Russell and Moran, 2006), but also in other complex interactions protecting the host from invasion by pathogenic microorganisms (a process known as “colonization resistance”) and modulating the insect immune system. Microbiome seems therefore to act in aphids (and actually not only in insects) as a sort of ecological immunity or extended immune system being able of affecting the efficiency of the host immune system and limiting the accumulation of pathobionts. As evident in the photo from a Nancy Moran paper, aphid bacteriocytes possess several symbiotic bacteria including the larger Buchnera primary symbionts (arrowheads) and the smaller H. defensa secondary symbionts (arrows).

In a recent paper, Chiu et al. (2012) reported a low survival of nymphs of the aphid Myzus varians at high temperatures as a consequence of the elimination of endosymbionts, such as Buchnera. This effect probably results from a temperature-mediated decrease in aphid endosymbionts, which synthesize amino acids essential for their insect hosts. In the last years, different roles have been suggested for symbionts other than the synthesis of amino acids only (Russell and Moran, 2006). Buchnera might, for instance, play a key role in aphid thermal tolerance since endosymbionts code for heat shock proteins, which deter degradation of host protein secondary structure (Dunbar et al., 2007). Secondary endosymbionts, such as Serratia simbiotica, play a similar role in the thermal tolerance of their host strengthening the ability of aphids to evolve further adaptations to overcome the impacts of warming (Russell and Moran, 2006).

Buchnera are at least partly able to survive at high temperatures because of constitutive expression of genes that are normally up-regulated in response to heat and aphids could be able to thrive under temperatures as high as 35°C in the laboratory (Dunbar et al., 2007). Surprisingly, a single nucleotide deletion in the Buchnera ibpA gene encoding for a small heat-shock protein virtually eliminates the transcriptional response of ibpA to heat stress and lowers its expression even at cool or moderate temperatures (Dunbar et al., 2007). In the present of this mutant allele, a short heat exposure in juveniles has strong effects on aphids that produce few or no progeny and contain almost no Buchnera, in contrast to aphids bearing symbionts without the deletion.

The ibpA mutated allele has appreciable frequencies in field populations supporting the view that lowering of ibpA expression improves host fitness under some conditions (Dunbar et al., 2007). As previously suggested, the response to stress (including thermal stress) is part of a large trade-off that related stress response to reproduction and immunity. This mutation by switching off the response to heat stimuli could favor aphid reproduction and immunity. However, the prolonged permanence of aphids at high temperatures (for instance in hot summer with daily mean temperature of 32.5 °C) results in the elimination of Buchnera reducing not only the thermal tolerance of aphids, but also their fecundity since the lack of endosymbionts results in a lost synthesis of amino acids essential for the hosts (Chiu et al., 2012).

According to these results, global warming could be difficultly faced by aphids in tropical regions due to Buchnera symbiont depletion.  Interestingly, in the presence of low density of primary symbionts, secondary symbionts (such as Hamiltonella defensa,  Serratia symbiotica, Regiella insecticola) could be more present affecting not only the aphid thermal tolerance to high temperatures clearly suggesting, but also their immune response due to symbionts (Poirié and Coustau, 2011).

The effects of global warming on the composition of aphid microbiota are of particular interest since, as recently reviewed by Poirié and Coustau (2011), the immune deficiency (IMD) signalling pathway was apparently non functional in aphids and no genes coding for peptidoglycan recognition proteins (PGRPs) and several well-conserved antimicrobial peptides, such as defensins and cecropins, have been predicted in the pea aphid Acyrthosiphon pisum genome (Gerardo et al., 2010), making the microbiota-based immunity essential to protect the host against natural enemies (Poirié and Coustau, 2011).

Even if effective, the symbiont-associated immunity appears to be more ephemeral and less stable than genetic resistance (Poirié and Coustau, 2011). Indeed, the rate of vertical transmission of symbionts is not always 100 %, so that bacteria can be lost, and their presence seems to be more energetically costly (Poirié and Coustau, 2011).

References

Chiu, M., Chen, Y., & Kuo, M. (2012). The effect of experimental warming on a low-latitude aphid, Myzus varians Entomologia Experimentalis et Applicata, 142 (3), 216-222 DOI: 10.1111/j.1570-7458.2011.01213.x

Dunbar, H.E., Wilson, A.C., Ferguson, N.R. & Moran, N.A. (2007). Aphid thermal tolerance is governed by a point mutation in bacterial symbionts. PLoS biology, 5 (5) PMID: 17425405

Poirié, M. & Coustau, C. (2011). The evolutionary ecology of aphids’ immunity. Inv. Surv. J., 8, 247-255

Russell, J.A & Moran, N.A. (2006). Costs and benefits of symbiont infection in aphids: variation among symbionts and across temperatures. Proceedings. Biological sciences / The Royal Society, 273 (1586), 603-10 PMID: 16537132

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Do you have endosymbionts useful for thermal tolerance?

One of the most studied symbioses occurs between aphids and the protobacterium Buchnera aphidicola that has been identified as primary endosymbiont of the pea aphid Acyrthosiphon pisum.
In view of its long association with aphids, Buchnera lost several genes, including those for the anaerobic respiration, the synthesis of different biomolecules (including amino sugars and some carbohydrates) and the biosynthesis of cell-surface components, such as lipopolysaccharides and phospholipids indicating that Buchnera is completely symbiotic. Buchnera also lost regulatory factors, so that it continuously overproduces tryptophan and other amino acids that are useful for aphids.

Figure. As reported in Nature, different pathways lost genes in the Buchnera genome. Pathways or steps for which no enzymes were identified are pink.

Almost all aphid species have 60–80 huge cells, called bacteriocytes, where Buchnera bacteria (in green in the figure) live. As reported for other symbionts, Buchnera is maternally transmitted to eggs and embryos through host generations, and the mutualism between the host and the bacteria is so obligate that neither can reproduce independently.
Different roles have been suggested for symbionts other than the synthesis of amino acids only. Buchnera might, for instance, play a key role in aphid thermal tolerance. Thermal tolerance of the primary endosymbiont Buchnera is attributed to genes coding for heat shock proteins, which deter degradation of protein secondary structure. Secondary endosymbionts (other optional bacterial species that can be present within the aphid body), such as Serratia simbiotica, play a similar role in the thermal tolerance of their host strengthening the ability of aphids to evolve further adaptations to overcome the impacts of warming.
Buchnera are at least partly able to survive at high temperatures because of constitutive expression of genes that are normally upregulated in response to heat and aphids could be able to thrive under temperatures as high as 35°C in the laboratory.

Unfortunately, the prolonged permanence of aphids at high temperatures (for instance in hot summer with daily mean temperature of 32.5 °C) results in the elimination of Buchnera reducing not only the thermal tolerance of aphids, but also their fecundity since the lack of endosymbionts results in a lost synthesis of amino acids essential for the hosts.

According to these results, global warming could be difficultly faced by aphids in tropical regions due to Buchnera symbiont depletion.  Interestingly, in the presence of low density of primary symbionts, secondary symbionts (such as Hamiltonella defensa,  Serratia symbiotica, Regiella insecticola) could be more present and improve aphid thermal tolerance to high temperatures clearly suggesting a role for heritable symbionts in the adaptation of aphids to their abiotic environments.

This is way I love evolution… there isn’t a unique solution to problems!

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